It is well known that cryopreservation of unfertilised human ooccytes is a technique which offers several advantages, especially whenever oocytes are to be preserved of patients who are at risk of ovarian hyperstimulation and can not transfer embryos during the in vitro fertilization treatment. However cryopreservation of oocytes, especially of fresh human oocytes, has run into greater technical difficulties than preservation of male gametes or embryos because of oocytes cytological peculiarity.
The low number of births from human cryopreserved oocytes, reported in literature, shows the technical difficulties of cryopreserving oocytes. Up to now researches carried out on oocytes cryopreservation provide congesting results regarding the more suitable and less damaging methods for maintaining cellular integrity and for getting a higher rate of viable oocytes.
As yet, a definitive protocol for cryopreserving human oocytes has not been established and the number of oocytes utilised up to now is still too low to determine a definitive methodology to be applied.
Human oocytes survival rate in cryopreservation depends, as well as on oocytes size, also on cryoprotectant used (composition, concentration and exposure time) and on freezing/thawing rate.
In the cryopreservation process, oocytes size is a very important parameter affecting the survival rate because the large quantity of water in ooplasm causes intracellular ice formation during the freezing, process: intracellular ice is one of the main responsible factors for intracellular structure damages.
Oocytes cryopreservation protocols usually include the following steps:    a) initially exposing the oocytes to a solution including a permeating cryoprotectant (e.g. 1,2-propanediol (PROH)), which aim is to reduce to a minimum intracellular structure damages caused by water crystallization;    b) subsequently exposing for a time of 2–5 min. the oocytes to a so-called loading solution including a mixture of a permeating cryoprotectant and a non permeating cryoprotectant (e.g. sucrose) to increase oocytes dehydration;    c) slowly cooling to −150° C.;    d) storing in liquid nitrogen (−196° C.);    e) thawing    f) diluting ad removing the cryoprotectants by exposure to so-called thawing solutions and returning to the physiological environment for further manipulations.
Cryoprotectants benefits are related to    I) their concentration,    II) exposure times,    III) the temperature at which they are added to oocytes.
Known methods for cryopreserving oocytes provide for using loading/thawing solutions including sucrose at a concentration of 0.1M or 0.2M.